Carotenoids are natural pigments widely present in the natural world. They are polyene pigments having a color in the range of yellow to red or purple. Astaxanthin is one type of naturally-occurring carotenoid that exists in a free state or as an ester, or as various types of pigment proteins upon binding with proteins.
Astaxanthin is widely used as a coloring agent for fish and for chicken eggs. Astaxanthin is also approved as a food additive and is widely used in fat and oil-processed foods, protein foods, aqueous liquid foods, etc. Furthermore, astaxanthin has an antioxidant activity against peroxidation of lipids induced by free radicals, a singlet oxygen-scavenging action which is several hundred times stronger than that of α-tocopherol, or the like; therefore, astaxanthin is expected to be used in functional foods, cosmetic products, or pharmaceuticals by utilizing the strong antioxidant activity.
Astaxanthin is distributed widely in the natural world, for example, in fishes such as salmon, trout and red sea bream; and crustaceans such as crab, shrimp, and krill. Astaxanthin is also produced by microorganisms such as bacteria belonging to the genera Agrobacterium, Brevibacterium, Paracoccus, Brevundimonas, Erythrobacter; the genus Haematococcus green algae; and the genus Phaffia yeasts. Carotenoids such as astaxanthin, zeaxanthin or the like are industrially produced by a chemical synthesis method, but naturally-derived carotenoids are desired in terms of safety.
With such a background, many methods for producing carotenoids containing astaxanthin especially derived from microorganisms such as green algae or yeasts, which are considered to be suitable for mass production, have been reported. However, since green algae and yeasts have low carotenoid productivity and have a strong cell wall, carotenoid extraction from their cultures is difficult.
On the other hand, bacteria belonging to the genus Paracoccus have advantages such as having a high carotenoid productivity, having great proliferation speed, and easy carotenoid extraction.
The E-396 strain (FERM BP-4283: deposited on Apr. 27, 1993 (date of original deposition), International Patent Organism Depositary, National Institute of Advanced Industrial Science and Technology, (Chuoh 6, Higashi 1-1-1, Tsukuba City, Ibaraki Prefecture, Japan)) is known as an example of an astaxanthin-producing bacterial strain belonging to the genus Paracoccus (Patent Document 1). Reported methods for obtaining a purified carotenoid-containing composition from this bacterial strain include: a method which performs extraction by contacting the bacterial cells with a cyclic hydrophilic organic compound, which is not favorable for use in production of food or such in terms of safety (Patent Document 2); a method using supercritical fluid extraction (Patent Document 3); a method in which bacterial cells are contacted with a water-soluble organic solvent, a non-polar solvent, and water to perform liquid-liquid extraction (Patent Document 4); and also a method of extraction by contacting bacterial cells with lower alcohols, then washing precipitates obtained by concentrating the extract using water-containing lower alcohols (Patent Documents 5 and 6).